The present invention generally relates to an immunological agglutination analysis and more particularly to a technique for testing or judging the existence of particular antigens or antibodies due to a particle agglutination reaction.
Heretofore, there have been proposed various types of such immunological agglutination judging methods for determining the blood types and various kinds of antigens and antibodies by detecting photoelectrically an agglutinated particle pattern or a non-agglutinated particle pattern formed by descending particles on a conically inclined bottom surface of a reaction vessel, while the reaction vessel is maintained substantially stationary. It has been further proposed to provide a number of reaction vessels each having a conical bottom surface in a matrix form in a base plate to constitute a microplate. In such an immunological agglutination testing method using the reaction vessel having the inclined bottom surface, when there is the agglutination reaction, the agglutinated particles are deposited on the inclined surface just like snow to form an uniformly deposited pattern, while in case of the non-agglutination reaction, the particles roll down along the inclined surface and are collected at the lowermost center portion, i.e. an apex of the cone to form a collected pattern. In one known method for detecting photoelectrically the uniformly deposited pattern and the collected pattern to distinguish therebetween, a light flux transmitted through the lowermost portion of the inclined surface is received by a light receiving element to produce a photoelectrically converted signal and then this signal is compared with a predetermined reference level to detect the existence or non-existence of the particle agglutination reaction. In another known method, an image of the particle pattern on the conical bottom surface is formed on an image plane on which two light receiving elements are arranged in such a manner that the first element can selectively receive an image on the lowermost portion of the conical bottom surface and the second element can exclusively receive an image on a portion of the inclined bottom surface surrounding said lowermost portion, and a difference between output signals of these elements is compared with a predetermined reference level to detect the existence of the particle agglutination reaction.
In the known methods, when the concentration of a particle suspension and an amount of the particle suspension delivered into the reaction vessel are varied, the outputs of the light detectors are also varied and thus, an accurate detection can not be effected. For instance, in case of determining the blood types, a sample of whole blood is first centrifuged to form a blood cell sediment and a given amount of the sedimented blood cells is sucked and delivered into the reaction vessel. In this case, the degree of sedimentation of blood cells during centrifugation might be affected by a variation of various conditions of the centrifuge, mechanical shock and vibration to which the sedimented blood cells are subjected after centrifugation and differences in viscosity and specific gravity of blood cells and blood plasma. Therefore, even if a predetermined amount of blood cell sediment is delivered, a variation in the amount and/or concentration of delivered blood cells might be produced. Further, such a variation might be produced due to a fluctuation of the delivery mechanism.
When the particle concentration and/or the amount of the delivered particle sample are varied, in the former method, the uniformly deposited particle pattern might be erroneously judged as the collected particle pattern when the particle concentration is too high, because the amount of light flux transmitted through the lowermost central portion of the conical bottom surface becomes too small, while the collected pattern might be judged as the uniformly deposited pattern when the particle concentration is too low, because the amount of light flux transmitted through the lowermost central portion is increased. Further, in the latter known method, the uniformly deposited pattern might be erroneously determined as a halfly deposited pattern which is produced by particles having a weak agglutination reaction, when the particle concentration is too high, because a part of particles which could not be fully agglutinated fall down into the lowermost portion. Contrary to this, when the particle concentration is too low, the halfly deposited pattern might be misjudged as the uniformly deposited pattern, and the collected pattern might be erroneously judged as the uniformly deposited pattern, because the amount of particles deposited at the lowermost portion becomes too small. Moreover, when the particle concentration becomes extremely small, even in case of the non-agglutination reaction, since an amount of particles collected in the lowermost portion becomes very small, the collected pattern could hardly be distinguished from the uniformly deposited pattern.